title: Reverse transcription-PCR analysis of the regulation of the manganese peroxidase gene family.
abstract: Manganese peroxidase (MnP) gene expression in the lignin-degrading fungus Phanerochaete chrysosporium is regulated by nutrient nitrogen levels and by Mn(II), the substrate for the enzyme, as well as by heat shock and other factors. Reverse transcription-PCR (RT-PCR) of total RNA can distinguish the mRNAs of each of the three sequenced P. chrysosporium mnp genes, i.e., mnp1, mnp2, and mnp3. Quantitative RT-PCR demonstrates that each of the three transcripts is present at a similar low basal level in nitrogen-sufficient cultures, with or without Mn, and in nitrogen-limited cultures lacking Mn. However, in 5-day-old, nitrogen-limited, stationary cultures supplemented with 180 microM Mn, the levels of the mnp1 and mnp2 transcripts increased approximately 100- and 1,700-fold, respectively, over basal levels. In contrast, under these conditions, the level of the mnp3 transcript did not increase significantly over the basal level. Quantitative RT-PCR of total RNA extracted from nitrogen-deficient, Mn-supplemented cultures on days 2 through 7 demonstrates that whereas the mnp1 transcript was present at relatively low levels on days 3 through 7, the mnp2 transcript level peaked on day 5 and the mnp3 transcript level peaked on day 3. Comparison of total RNA extracted on day 5 from nitrogen-deficient, Mn-supplemented stationary and agitated cultures indicates that in stationary cultures, mnp2 was the major expressed mnp gene, whereas in large agitated cultures, mnp1 was the major expressed mnp gene.
authors.affiliation: Department of Biochemistry and Molecular Biology, Oregon Graduate Institute of Science and Technology, Portland 97291-1000, USA.
authors.forename: J M
authors.initials: JM
authors.lastname: Gettemy
authors.forename: B
authors.initials: B
authors.lastname: Ma
authors.forename: M
authors.initials: M
authors.lastname: Alic
authors.forename: M H
authors.initials: MH
authors.lastname: Gold
mesh_terms: D001487:Basidiomycota / Q000235:genetics*; D005800:Genes, Fungal; D010544:Peroxidases / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis
chemical_list: D012333:RNA, Messenger; D010544:Peroxidases; C051129:manganese peroxidase